Opioids such as morphine are among the most important tools used to treat pain, however their use is limited due to the development of tolerance and other negative side effects. Cannabinoids such as 9- tetrahydracannabinol (THC) are also used for pain relief, however their low efficacy and side effects also limit their clinical utility. Opioids and cannabinoids interact in several ways to produce better pain relief than either drug alone. Recent clinical data in humans demonstrates that cannabinoid administration can improve pain relief in patients already taking opioids for chronic pain. Our preliminary data supports these findings, and suggests that this effect is at least partially mediated by the midbrain periaqueductal gray (PAG). The PAG is critically involved in descending pain modulation and the development of morphine tolerance. The goal of the proposed studies is to elucidate the cellular adaptations in cannabinoid signaling and neurotransmission that occur in the PAG after the development of opioid tolerance. The following two Specific Aims will address these goals, using in vitro electrophysiology techniques: 1) Characterize the changes in cannabinoid-mediated synaptic transmission after chronic morphine exposure. 2) Determine the cellular mechanisms underlying morphine tolerance-induced changes in cannabinoid modulation of neurotransmission in the PAG. The experiments in Aims 1 and 2 will employ whole-cell patch clamp techniques to measure changes in synaptic transmission after chronic opioid exposure, and to determine the molecular mechanisms underlying these changes. Behavioural testing in rats will also be employed to assess antinociception, and verify the development of morphine tolerance. The proposed studies will identify the molecular mechanisms underlying the changes that occur following chronic opioid treatment, in a brain region that is critically involved in opioid tolerance. The field of opioid/cannabinoid interaction is still developing, and characterizing the physiology of this system will have a significant impact on the field. Full understanding of this interaction is critial to our long-term goal of identifying cannabinoid-related cellular targets that may be used to overcome tolerance and improve the analgesic efficacy of opioids. Such an innovation could lead to a major advance in clinical pain management.